Molecular Characterization of the PmrA Regulon*

The two-component system PmrA/PmrB ofSalmonella enterica controls expression of several loci including those mediating modifications in the lipopolysaccharide that result in polymyxin resistance. To gain insight in the regulation of polymyxin resistance, we mapped the transcription start sites of the PmrA-regulated genes pmrC, pmrG, pbgPE, andugd and identified a conserved sequence in the promoter region of the first three genes. His-tagged PmrA protein could gel shift DNA fragments containing the promoters of the pmrC,pmrG, and pbgPE genes but not theudg promoter. DNase I footprinting analysis of thepmrC, pmrG, and pbgPE promoters indicate that phosphorylated as well as unphosphorylated PmrA bind to a 16-base pair imperfect inverted repeat sequence (5′-TTAAKTTCTTAAKGTT-3′), which is found 40, 80, and 38 nucleotides upstream from the transcription start sites of the pmrC, pmrG, and pbgPE genes, respectively. Our data suggest that a PmrA dimer activates transcription of the divergentpmrG and pbgPE promoters by binding to a single site in the pmrG-pbgPE intergenic region and that the ugd gene is regulated by the PmrA/PmrB system only indirectly.

[1]  W. Liu,et al.  Bacillus subtilis PhoP binds to the phoB tandem promoter exclusively within the phosphate starvation-inducible promoter , 1997, Journal of bacteriology.

[2]  W. Shafer,et al.  Late intraphagosomal hydrogen ion concentration favors the in vitro antimicrobial capacity of a 37-kilodalton cationic granule protein of human neutrophil granulocytes , 1986, Infection and immunity.

[3]  K. Murakami,et al.  Nature of DNA binding and RNA polymerase interaction of the Bordetella pertussis BvgA transcriptional activator at the fha promoter , 1997, Journal of bacteriology.

[4]  R. Gourse,et al.  A third recognition element in bacterial promoters: DNA binding by the alpha subunit of RNA polymerase. , 1993, Science.

[5]  S. Miller,et al.  PhoP-PhoQ activates transcription of pmrAB, encoding a two-component regulatory system involved in Salmonella typhimurium antimicrobial peptide resistance , 1996, Journal of bacteriology.

[6]  J. Stock,et al.  Analysis of two-component signal transduction systems involved in transcriptional regulation. , 1996, Methods in enzymology.

[7]  E. Groisman,et al.  Two-component regulatory systems can interact to process multiple environmental signals , 1996, Journal of bacteriology.

[8]  W. Shafer,et al.  Lipid A and resistance of Salmonella typhimurium to antimicrobial granule proteins of human neutrophil granulocytes , 1984, Infection and immunity.

[9]  A. Ninfa,et al.  Protein phosphorylation and regulation of adaptive responses in bacteria. , 1989, Microbiological reviews.

[10]  K. Makino,et al.  Role of the sigma 70 subunit of Escherichia coli RNA polymerase in transcription activation. , 1994, Journal of molecular biology.

[11]  J. Gunn,et al.  The Salmonella typhi melittin resistance gene pqaB affects intracellular growth in PMA-differentiated U937 cells, polymyxin B resistance and lipopolysaccharide. , 1999, Microbiology.

[12]  E. Groisman,et al.  Regulation of polymyxin resistance and adaptation to low-Mg2+ environments , 1997, Journal of bacteriology.

[13]  A. Cheung,et al.  Molecular Interactions between Two Global Regulators,sar and agr, in Staphylococcus aureus * , 1998, The Journal of Biological Chemistry.

[14]  J. Demoss,et al.  NarL‐phosphate must bind to multiple upstream sites to activate transcription the narG promoter of Escherichia coli , 1994, Molecular microbiology.

[15]  T. Kawamoto,et al.  Signal transduction in the phosphate regulon of Escherichia coli involves phosphotransfer between PhoR and PhoB proteins. , 1989, Journal of molecular biology.

[16]  K. Roland,et al.  Isolation and characterization of a gene, pmrD, from Salmonella typhimurium that confers resistance to polymyxin when expressed in multiple copies , 1994, Journal of bacteriology.

[17]  M. D. Snavely,et al.  Magnesium transport in Salmonella typhimurium. Regulation of mgtA and mgtB expression. , 1991, The Journal of biological chemistry.

[18]  P. Helenamakela.  Isolation and genetic characterization of polymyxin-resistant mutants of Salmonella , 1978 .

[19]  J. S. Parkinson,et al.  Communication modules in bacterial signaling proteins. , 1992, Annual review of genetics.

[20]  S. Miller,et al.  PmrA–PmrB‐regulated genes necessary for 4‐aminoarabinose lipid A modification and polymyxin resistance , 1998, Molecular microbiology.

[21]  T. Mizuno,et al.  Phosphorylation of a bacterial activator protein, OmpR, by a protein kinase, EnvZ, results in stimulation of its DNA-binding ability. , 1989, Journal of biochemistry.

[22]  D. Dubnau,et al.  ComA, a phosphorylated response regulator protein of Bacillus subtilis, binds to the promoter region of srfA , 1993, Journal of bacteriology.

[23]  K. Roland,et al.  Spontaneous pmrA mutants of Salmonella typhimurium LT2 define a new two-component regulatory system with a possible role in virulence , 1993, Journal of bacteriology.

[24]  E. Groisman,et al.  Mg2+ as an Extracellular Signal: Environmental Regulation of Salmonella Virulence , 1996, Cell.

[25]  M. Schwartz,et al.  Positive control of transcription initiation in bacteria. , 1984, Annual review of genetics.

[26]  Richard H. Ebright,et al.  Promoter structure, promoter recognition, and transcription activation in prokaryotes , 1994, Cell.

[27]  I. Sutherland.  Surface carbohydrates of the prokaryotic cell , 1977 .

[28]  I. Kilpeläinen,et al.  Increased substitution of phosphate groups in lipopolysaccharides and lipid A of the polymyxin‐resistant pmrA mutants of Salmonella typhimurium: a 31P‐NMR study , 1994, Molecular microbiology.

[29]  N. W. Davis,et al.  The complete genome sequence of Escherichia coli K-12. , 1997, Science.

[30]  C. Haidaris,et al.  Mutants of Salmonella typhimurium that cannot survive within the macrophage are avirulent. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[31]  R. Kadner,et al.  Protein Phosphorylation Affects Binding of the Escherichia coli Transcription Activator UhpA to the uhpT Promoter* , 1997, The Journal of Biological Chemistry.

[32]  V. Weiss,et al.  A common switch in activation of the response regulators NtrC and PhoB: phosphorylation induces dimerization of the receiver modules. , 1995, The EMBO journal.